Project Summary Abstract Mounting mechanistic and translational studies support the use of cell-based therapies to repair myocardial tissue destroyed by infarction and to restore cardiac function. Several phenotypically distinct subsets of adult primitive cell populations have been shown to improve cardiac structure and function in animal models of myocardial infarction (MI) and heart failure. Small clinical trials of stem cell therapy have recapitulated these beneficial effects in patients with ischemic cardiomyopathy. Recent discovery of cardiac stem cells (CSCs) has sparked intense hope for the development of promising stem cell therapies for cardiac repair/regeneration because CSCs are inherently programmed to reconstitute cardiac tissue. In recent studies, we found that intracoronary delivery of CSCs to rats with either acute or chronic MI and to pigs with chronic MI ameliorated cardiac function and regenerated new cardiac cells. However, human patients needing cardiac reparative therapies generally possess an array of cardiovascular risk factors such as hypercholesterolemia (HC), diabetes, hypertension etc. With the recent surge of interest in cell therapies for patients, it is important to understand the impact of these risk factors on cell-mediated cardiac repair. In particular, HC is a highly prevalent risk factor and contributes to a range of pathophysiological consequences. Hence, the overall goal of this proposal is to investigate the impact of HC on CSC-mediated cardiac repair. Our fundamental hypothesis is that depending on the specific conditions, cholesterol can be beneficial or detrimental in CSC-mediated cardiac repair. We propose that mild elevations of plasma cholesterol or the presence of the minimally oxidatively modified form of LDL-cholesterol precondition both the myocardium and the CSCs; the resulting combination of a primed myocardial microenvironment for cell engraftment and enhanced paracrine signaling mechanisms of preconditioned CSCs work in concert to enhance CSC-mediated cardiac repair. We further propose that marked elevations of plasma cholesterol or the presence of the completely oxidized form of LDL- cholesterol provoke oxidative injury to both the CSCs and the myocardium, leading to loss of efficacy of cell therapies for cardiac repair. We will test these hypotheses under 3 specific aims using both in vitro cultured CSCs and cardiomyocytes in the presence of differently modified LDLs and in vivo rat models of MI with different levels of plasma cholesterol. Aim 1 will determine the effects of HC on CSC-mediated cardiac repair in vivo; Aim 2 will determine the effects of LDLs on CSC function and reparative capability in vitro; and Aim 3 will determine whether pretreatment of CSCs with differently modified LDLs in vitro alters the efficacy of cardiac reparative therapy in vivo. Given that plasma cholesterol is a modifiable risk factor, but is also essential for cellular function, understanding the effects of this prevalent risk factor on stem cell-based therapies will have translational and mechanistic importance. This project will provide novel insights into a much-needed preclinical framework to develop cell-based therapies for cardiac repair in patients with cardiovascular risk factors.